Published online Aug 26, 2020. doi: 10.4252/wjsc.v12.i8.841
Peer-review started: February 25, 2020
First decision: May 26, 2020
Revised: July 17, 2020
Accepted: August 1, 2020
Article in press: August 1, 2020
Published online: August 26, 2020
Cigarette smoking (CS) is the most common method of consuming tobacco. Deleterious effects on bone integrity, increased incidence of fractures, and delayed fracture healing are all associated with CS. Over 150 of the 6500 molecular species contained in cigarette smoke and identified as toxic compounds are inhaled by CS and, via the bloodstream, reach the skeletal system. New technologies designed to develop a reduced-risk alternative for smokers are based on electronic nicotine delivery systems, such as e-cigarettes and tobacco heating systems (THS). THS are designed to heat tobacco instead of burning it, thereby reducing the levels of harmful toxic compounds released.
To examine the effects of THS on osteoprogenitor cell viability and function compared to conventional CS.
Human immortalized mesenchymal stem cells (n = 3) and primary human pre-osteoblasts isolated from cancellous bone samples from BG Unfall Klinik Tübingen (n = 5) were osteogenically differentiated in vitro with aqueous extracts generated from either the THS 2.4 “IQOS” or conventional “Marlboro” cigarettes for up to 21 d. Cell viability was analyzed using resazurin conversion assay (mitochondrial activity) and calcein-AM staining (esterase activity). Osteogenic differentiation and bone cell function were evaluated using alkaline phosphatase (AP) activity, while matrix formation was analyzed through alizarin red staining. Primary cilia structure was examined by acetylated α-tubulin immunofluorescent staining. Free radical production was evaluated with 2’,7’-dichlorofluorescein-diacetate assay.
Our data clearly show that THS is significantly less toxic to bone cells than CS when analyzed by mitochondrial and esterase activity (P < 0.001). No significant differences in cytotoxicity between the diverse flavors of THS were observed. Harmful effects from THS on bone cell function were observed only at very high, non-physiological concentrations. In contrast, extracts from conventional cigarettes significantly reduced the AP activity (by two-fold) and matrix mineralization (four-fold) at low concentrations. Additionally, morphologic analysis of primary cilia revealed no significant changes in the length of the organelle involved in osteogenesis of osteoprogenitor cells, nor in the number of ciliated cells following THS treatment. Assessment of free radical production demonstrated that THS induced significantly less oxidative stress than conventional CS in osteoprogenitor cells.
THS was significantly less harmful to osteoprogenitor cells during osteogenesis than conventional CS. Additional studies are required to confirm whether THS is a better alternative for smokers to improve delays in bone healing following fracture.
Core tip: Aqueous extracts (AE) generated with tobacco heating systems (THS) showed no differences in suspended particles compared to the cell culture medium. This finding supports previous studies demonstrating reduced levels of harmful constituents reported in THS AE in comparison to conventional cigarettes AE. The time to consume one unit (stick/cigarette) was longer for THS than for a conventional cigarette. The pH from both AE fractions was similar to the cell culture medium. Following a single exposure, THS AE was significantly less toxic to bone-forming cells and osteoprogenitor cells than conventional cigarettes AE. The cytotoxicity observed following THS exposure was associated with very high, non-physiological concentrations. No significant differences in cytotoxicity were observed between different flavors of THS AE. Moreover, THS AE displayed less impact on osteogenic differentiation of osteoprogenitor cells and the function of bone-forming cells when compared to conventional cigarettes AE. Finally, compared to conventional cigarettes AE, THS AE induced lower levels of oxidative stress due to the reduced level of harmful constituents, resulting in less damage to primary cilia structure and reduced impact on osteogenic differentiation.